Professional Documents
Culture Documents
Biosafety
Biosafety
INDEX Introduction Risk Assessment Biosafety Recommendations and Containment Measures Precautionary measures for handling cell cultures
Introduction
The use of animal and human cell cultures has become very beneficial for diverse applications in the fields of biotechnology, medicine and veterinary. Originally used as substrates for the production of viral vaccines (Salk polio vaccine on monkey kidney cells, rabies, mumps and rubella vaccine using the WI-38 cell line), animal and human cell cultures became an indispensable tool to study intra- or intercellular responses and to serve as in vitro model for research. It has also been used for in vitro diagnosis of viruses or for the production of a wide range of biological products (hormones, interleukins, interferons and growth factors), including potential diagnostic and therapeutic products. More recently, cellular entity features associated to human and mammalian cell cultures have also gained interest in the search of new therapeutic approaches such as allo-/xeno- transplantation or cell-based gene therapy. Animal and human cell cultures in vitro, biosafety concerns have pointed to the risks with respect to human (as well as animal or plant) health and environmental considerations. A maximal reduction of these risks necessitates a thorough risk assessment of the given cell cultures, taking into account the type of manipulation and the implementation of appropriate containment measures. Depending on the purpose or the type of activity, the use of animal cell cultures may fall within the scope of several regulatory provisions. In Europe, for example, as the manipulation of animal cell cultures may pose a risk related to the exposure of the worker to biological agents, this type of activity is covered by the European Directive 2000/54/EC. In many cases, tissue culture work will also involve the use of genetically-modified cell lines, in which case a risk assessment should be made in accordance with the provisions of the Directive 98/81/EC related to the contained use of genetically-modified organisms. Moreover, cell culturing activities aiming at manufacturing biopharmaceuticals are covered by the Regulation (EC) No 726/2004 laying down procedures for the authorization and supervision of medicinal products for human and veterinary
use, whereas activities that involve the use of human cells and tissues for application to the human body fall within the scope of the Directive 2004/23/EC which provides standards for the use of human cells and tissues in the category of cell therapy. Biosafety recommendations are principally aimed at providing maximal protection of human health and environment, it is recognized that many of the precautionary measures would directly benefit the quality of research activities involving animal cell cultures as cross-contamination or inadvertent contamination with biologic agents are plaguing many researchers, often leading to inaccurate data, misinterpretation of results and a considerable waste of time and energy.
Risk Assessment
Risk assessment of animal cell cultures is based on both the intrinsic properties of the cell cultureincluding subsequent properties acquired as a result of genetic modificationand the possibility that the cell culture may inadvertently or deliberately become contaminated with pathogens. In addition, the risks encountered during handling of animal cell cultures should be evaluated with a careful consideration of the type of manipulation.
such as the existence of effective therapies or prophylaxis. An evaluation of these criteria has been used to classify pathogens into classes of biological risk, also called Risk Groups. The four risk categories range from Risk Group 1, where biologic agent is unlikely to cause human disease, to Risk Group 4, where the agent causes severe human disease and present serious hazard to workers with a potential of spreading to the community. Contrary to agents of lower Risk Groups, there are usually no effective prophylaxes or treatments available for Group 4 Biological Agents.
assessment is to define biological hazards in order to be able to eliminate or prevent risks to both human health and the environment.
BIOSAFETY CABINETS
Biosafety cabinets are used to provide primary containment in the laboratory when the investigator is using potentially infectious materials. There are three types of biological safety cabinets: Class I: The Class I biological safety cabinet is an open-front negative pressure cabinet The exhaust air from the cabinet is filtered by a high-efficiency particulate air (HEPA) filter. The Class I biosafety cabinet will provide personnel and environmental protection, but not product protection. Class II: The Class II vertical laminar-flow biological cabinet is an open-front, ventilated cabinet. This cabinet provides a HEPA-filtered, recirculated mass airflow within the work space. The exhaust air from the cabinet is also filtered by HEPA filters. Thus, the Class II biosafety cabinet will provide personnel, environment and product protection. While HEPA filters are effective for trapping particulates and infectious agents, these filters will not capture volatile chemicals or gases Class III: The Class III cabinet is a totally enclosed ventilated cabinet of gas-tight construction. Operations within the Class III cabinet are conducted through attached rubber gloves. When in use, the Class III cabinet is maintained through negative air pressure of at least 0.5 inches water gauge. Supply air is drawn into the cabinet through HEPA filters. The cabinet exhaust air is filtered by two HEPA filters, installed in series, before discharge outside of the facility. The exhaust fan for the Class III cabinet is generally separate from the exhaust fans of the facility's ventilation system. The use of a Class II cabinet in the microbiological laboratory offers the additional capability and advantage of protecting materials contained within it from extraneous airborne contaminants. This capability is provided by the HEPAfiltered, recirculated mass airflow within the workspace. Personnel protection provided by Class I and Class II cabinets is dependent on the inward airflow. Since the face velocities are similar, they
generally provide an equivalent level of personnel protection. The use of these cabinets alone, however, is not appropriate for containment of highest-risk infectious agents because aerosols may accidentally escape through the open front. When Class III cabinets are required, all procedures involving infectious agents (usually Classes 3, 4 or 5) are performed within them. The World Health Organization and regulatory authorities, both in United States and in Europe, have formulated guidelines designed to minimize any potential risk for transmission of infectious agents in case animal cell cultures are used for the industrial production of biological with therapeutic purposes (EMEA, FDA, ICH, WHO). It includes extensive testing of the cell banks, the unpurified bulk material, as well as the final product with particular attention for viral clearance processes. Hardly any guidance has been provided for the extent of detecting possible contaminants in case animal cell cultures are used for in vitro research or diagnostic activities or for purposes other than therapeutics or production of biopharmaceuticals. It is recognized that there is no single test suitable for detecting all possible contaminants. Therefore, the choice of detection technique depends on the contaminating pathogen and often a combination of methods is necessary to enhance detection in important samples such as master cell banks. Table gives an overview of the use, detection capability and limitations of assays that might be used to detect and/or identify adventitious agents.